The bacteriophage PRD1 is a lipid bearing phage that infects a wide variety of gram-negative bacteria, including Escherichia coli and Salmonella typhimurium when they harbor the appropriate plasmid. It contains a linear duplex DNA molecule that is covalently bound by its 5' ends to a terminal protein. Like adenovirus and the Bacillus phage φ29, PRD1 specifies its own DNA polymerase which is able to utilize the phage encoded terminal protein to prime DNA synthesis. In addition to these two proteins, PRD1 also specifies an additional replication protein (p12) of unknown function. We have sequenced the origins of replication (termini of the genome) as well as the right most 1700 bp of the bacteriophage PRD1 genome. The right most 1700 bp encompasses the right early region and completes the sequence of all PRD1 early functions. We report here that the PRD1 genome contains a perfect 111 bp inverted terminal repeat. Furthermore, statistical analyses of the right 1700 bp, as well as the examination of transcription and translation signals has allowed us to assign gene XII to an open reading frame and to infer the direction of both early and late transcription. Gene XII, which has been implicated in the replication process and the regulation of gene expression is predicted to encode a 16.7 Kdal protein. Data base searches have revealed a possible evolutionary relationship between this protein and the ε-subunit of E. coli DNA polymerase III. We have also mapped right, early transcription of the PRD1 genome. This has corroborated our inference concerning the direction of right early transcription and confirmed our assignment of gene XII to an open reading frame. It has also revealed that two putative rho-independent terminators are functional in vitro and that the putative right early promoter is utilized in vivo and in vitro. The data presented here have permitted us to ascertain the general genomic and transcriptional organization of PRD1 and to predict the primary structure of the product of gene XII. These results, in turn, have allowed us to develop hypotheses concerning the evolution of linear, protein primed DNA's and the function of gene XII.

The bacteriophage PRD1 is a lipid bearing phage that infects a wide variety of gram-negative bacteria, including Escherichia coli and Salmonella typhimurium when they harbor the appropriate plasmid. It contains a linear duplex DNA molecule that is covalently bound by its 5' ends to a terminal protein. Like adenovirus and the Bacillus phage φ29, PRD1 specifies its own DNA polymerase which is able to utilize the phage encoded terminal protein to prime DNA synthesis. In addition to these two proteins, PRD1 also specifies an additional replication protein (p12) of unknown function. We have sequenced the origins of replication (termini of the genome) as well as the right most 1700 bp of the bacteriophage PRD1 genome. The right most 1700 bp encompasses the right early region and completes the sequence of all PRD1 early functions. We report here that the PRD1 genome contains a perfect 111 bp inverted terminal repeat. Furthermore, statistical analyses of the right 1700 bp, as well as the examination of transcription and translation signals has allowed us to assign gene XII to an open reading frame and to infer the direction of both early and late transcription. Gene XII, which has been implicated in the replication process and the regulation of gene expression is predicted to encode a 16.7 Kdal protein. Data base searches have revealed a possible evolutionary relationship between this protein and the ε-subunit of E. coli DNA polymerase III. We have also mapped right, early transcription of the PRD1 genome. This has corroborated our inference concerning the direction of right early transcription and confirmed our assignment of gene XII to an open reading frame. It has also revealed that two putative rho-independent terminators are functional in vitro and that the putative right early promoter is utilized in vivo and in vitro. The data presented here have permitted us to ascertain the general genomic and transcriptional organization of PRD1 and to predict the primary structure of the product of gene XII. These results, in turn, have allowed us to develop hypotheses concerning the evolution of linear, protein primed DNA's and the function of gene XII.

en_US

dc.type

text

en_US

dc.type

Dissertation-Reproduction (electronic)

en_US

dc.subject

Bacteriophages -- Genetics

en_US

dc.subject

DNA polymerases -- Analysis

en_US

thesis.degree.name

Ph.D.

en_US

thesis.degree.level

doctoral

en_US

thesis.degree.discipline

Molecular and Cellular Biology

en_US

thesis.degree.discipline

Graduate College

en_US

thesis.degree.grantor

University of Arizona

en_US

dc.contributor.advisor

Ito, Junetsu

en_US

dc.contributor.committeemember

Bernstein, Dr. Harris

en_US

dc.contributor.committeemember

Duffy, Dr. John

en_US

dc.contributor.committeemember

Lindell, Dr. Thomas

en_US

dc.contributor.committeemember

Spizizen, Dr. John

en_US

dc.identifier.proquest

9013145

en_US

dc.identifier.oclc

703433973

en_US

All Items in UA Campus Repository are protected by copyright, with all rights reserved, unless otherwise indicated.